Although various techniques for saccharifying cellulose are available, the enzymatic saccharification technique, which requires less energy but produces a high yield of sugar, has been in the mainstream of development. Cellulase, which is a cellulose-degrading enzyme, is broadly divided into cellobiohydrolases, which act on the crystalline regions of cellulose (and may be referred to as “CBH” in this specification), and endoglucanases, which act inside the cellulose molecular chain to reduce the molecular weight. β-glucosidase acts on a hydrosoluble oligosaccharide or cellobiose to catalyze the hydrolysis of their β-glycosidic bonds. Of these, cellobiohydrolases are most important for efficient saccharification and are being used in great amounts because cellobiohydrolases act on the crystalline regions of cellulose to hydrolyze the cellulose from its end. CBH is divided into two types, CBHI and CBHII, based on the difference in action mechanism. CBHI, an enzyme classified into GH7, cleaves a cellulose chain from its reducing end into cellobiose units. CBHI has a tunnel structure with four long loops that cover the active site and substrate-binding site. A cellulose chain can pass through the tunnel and is cleaved from its end into cellobiose units. CBHII is an enzyme classified into GH6, and cleaves a cellulose chain from its non-reducing end into cellobiose units. CBHII has two loops covering the active site, and the catalytic site forms a tunnel structure because of the loops.
Saccharification of biomass using the group of enzymes needs to be more efficient. In particular, improving the thermal stability (heat resistance) of the enzymes will enable contamination prevention, increased reaction efficiency, and more efficient saccharification through recycling of the enzymes.